1) Rat anti-mouse monoclonal antibodies (MAb) which block or augment killing of rat tumor target cells by murine cytotoxic T lymphocytes (CTL) will be obtained. Whether MAb block at the stage of crawling activity, specific antigen recognition and adhesion, or delivery of the lethal hit will be determined. Analogous mixed-lymphocyte reaction (MLR) blocking MAb will also be obtained. Antigen-negative mutant CTL clones will be selected by complement-mediated lysis or in the fluorescence-activated cell sorter (FACS). Antigens which are sites for blocking and mutations in which lead to loss of CTL activity are implicated in lytic mechanisms. The molecular weight and subunit structure of these molecules will be studied after radiolabeling and immunoprecipitation. Clonal variation suggestive of antigen receptors will be examined by peptide mapping. 2) Subclass-specific and broadly reactive IgG and IgM mouse MAb to rat IgG will be utilized in two-color immunofluorescence with rat MAb of specific subclasses, as screening reagents in indirect assays, and as facilitating IgM reagents for lysis with complement. 3) To obtain MAb to previously unrecognized antigens, membrane glycoproteins depleted of previously identified antigens with MAb immunoadsorbents will be used for immunization and/or coupled to fluorescent beads for selection and cloning of hybrid cells on the FACS. These and previously obtained MAb will be used to characterize antigen molecular structure after immunoprecipitation, cell distribution with the FACS and staining of thin sections, for correlation with other markers using two-color fluorescence, and for presence on helper, cytotoxic, and suppressor subpopulations after lysis or sorting. 4) An antigenic determinant found on spleen cell but not most thymocyte Thy-1 will be studied for distribution and protein or carbohydrate nature. Alpha-chains of Mac-1 and LFA-1 antigens will be compared for carbohydrate and tryptic peptide map differences, and for muturation or differentiation-related changes in expression in lymphocytes, granulocytes, and monocytes. This basic research on lymphocyte antigen structure and function should suggest many avenues for therapy and diagnosis in neoplasms and immune disorders.